1,4-Dichloro-2,2,5,5-tetrasubstituted-3,6-piperazinediones

ABSTRACT

There is provided, novel N-chloramine compounds of the 1,4dichloro-2,2,5,5-tetrasubstituted-3,6-piperazinedione type, having the formula:   WHERE R1, R2, R3, R4, which may be the same or different, each represent a member selected from the group consisting of an alkyl group of from 1 to 20 carbon atoms and an aryl group. The compounds encompassed by the above generic formula exhibit antibacterial activity, but most importantly, the compounds encompassed within the above generic formula exhibit enhanced stability over the closest related prior art compounds, when such compounds are employed in aqueous solution for antibacterial application.

United States Patent 1 Bodor et al.

[ 1,4-DICHLORO-2,2,5,5-

TETRASUBSTlTUTED-3,6-PIPERAZINED- [ONES [75} Inventors: Nicolae S. Bodor; James J.

Kaminski, both of Lawrence, Kans.

[73] Assignee: lNTERx Research Corporation,

Lawrence, Kansl [22] Filed: Apr. 1, 1974 [2]} Appl. No.: 456,743

[52] US. Cl. 260/268 DK; 424/250 [5 l] Int. Cl. C07D 51/72 [58] Field of Search 260/268 DK [56] References Cited UNITED STATES PATENTS 3.l36,598 6/l964 Kokorudz 260/268 DK 3 l42,530 7/l964 Kokorudz 260/268 DK Primary Examiner-Donald G. Daus Assistant Examiner.lose Tovar Attorney, Agent, or Firm-Charles N. Blitzer June 24, 1975 [57] ABSTRACT There is provided, novel N-chloramine compounds of the l,4-dichloro-2,2,5,5-tetrasubstituted-3 ,-piperazinedione type, having the formula:

6 Claims, No Drawings 1 l,4-DICHLORO-2,2,5 ,5-TETRASUBSTITUTED-3 ,6- PIPERAZINEDIONES BACKGROUND OF THE INVENTION 1. Field Of The Invention The present invention relates to certain novel N- chloramine compounds of the l,4-dichloro-2,2,5,5- tetrasubstituted-3,6-piperazinedione type and more particularly, the present invention relates to such compounds, which exhibit excellent antibacterial activity. However, most importantly, these compounds exhibit enhanced stability over the closest related prior art compounds in aqueous solution.

2. Description of the Prior Art US. Pat. No. 3,l36,598 to Kokorudz (hereinafter- Kokorudz") discloses certain l,4-dichloro2,5- diketopiperazine compounds, in which the carbon atom, adjacent to the carbonyl group can contain two hydrogen atoms or in the alternative, a hydrogen atom in combination with a methyl or an ethyl group. These compounds are useful as antibacterial agents; in other words, such compounds can be employed in aqueous solution for the purpose of disinfection. In addition, the compounds described in this patent, due to their high chlorine potential, can also serve as effective bleaching agents, and more specifically, bleaching agents for textile fabrics, such as cottons. (See, US. Pat. No. 3,l42,5 30.)

Applicants have extensively studied the compounds disclosed by Kokorudz and have observed that while these compounds do exhibit excellent antibacterial activity in aqueous solution, such compounds when contained in an aqueous solution, are highly unstable. That is, when such compounds are introduced into an aqueous solution for application as antibacterial agents, such solutions must be used almost immediately as the compounds disclosed by Kokorudz will rapidly decompose, thus precluding sufficient germicidal activity to be achieved.

SUMMARY OF THE INVENTION Accordingly, it is one object of the present invention to provide novel N-chloramine compounds of the l,4-

wherein R,,R R R which may be the same or different, each represent a member selected from the group consisting of an alkyl group of from 1 to 20 carbon atoms and an aryl group.

Illustrative alkyl groups are those of methyl, ethyl, propyl, butyl, pentyl, decyl, tetradecyl, hexadecyl, and eicosanyl and as illustrative aryl groups, phenyl, and naphthyl are exemplified. However, those alkyl groups containing 1-5 carbon atoms are preferred.

The unique stability exhibited by the compounds of the present invention as opposed to those compounds disclosed by Kokorudz results from the fact that the compounds of this invention do not contain any alpha hydrogen atoms on the carbon atom adjacent to the carbonyl group of the compounds disclosed herein.

During applicants investigation of those compounds disclosed by Kokorudz, they observed a relationship between stability and the lack of alpha-hydrogen atoms on the carbon atom adjacent to the carbonyl group in the piperazinedione nucleus. In other words, applicants observed that when all alpha-hydrogens on the carbon atoms adjacent the carbonyl groups of the piperazinedione nucleus were substituted with alkyl groups of from I to 20 atoms and/or aryl groups, stability of such resulting compounds was enhanced when compared to the stability exhibited by Kokorudzs compounds.

DETAILED DESCRIPTION OF THE INVENTION The compounds of the present invention can be prepared by a simple reaction scheme. Basically, these compounds are conveniently prepared by chlorinating the appropriate 2,2,5,5-tetrasubstituted-3,6-piperazinedione as illustrated below:

dichloro-2,2,5,itetrasubstituted-3,6-piperazinedione type, exhibiting sufficient antibacterial activity, ie germicidal and disinfecting activity.

Still, another object of the present invention is to provide novel N-chloramine compounds of the type described, where, in addition to such compounds exhibiting sufficient antibacterial activity, such compounds will exhibit a superior stability to those compounds described by Kokorudz.

Accordingly, the present invention is directed to the discovery of novel N-chloramine compounds of the l,- 4-dichloro-2,2,5,5tetrasubstituted-3,6-piperazinedione type, which have the formula:

ate dipeptide following conventional procedures as described by J. P. Greenstein and M. Winitz in the text entitled Chemistry of Amino Acids," Volume II, J. Wiley and Sons, Incorporated, New York, N.Y., l96l, Chapter ID.

The present invention will be better understood from a review of the following examples, which are simply illustrative and non-limitative of the present invention.

EXAMPLE 1 (Preparation of l,4-dichloro-2,2,5 ,5-tetramethyl-3 ,fi-piperazinedione) Firstly, the precursor compound (2,2,5,5- tetramethyl-3,o-piperazinedione) was prepared from aIpha-aminoisobutyric acid following the procedure described by S. M. McElvain and E. H. Pryde. THE JOURNAL OF THE AMERICAN CHEMICAL SOCI- ETY, 7l,326(1949). Reference to the general procedure used is made in the Greenstein and Winitz article referred to earlier in this Application.

Once the 2,2,5,5-tetramethyl3,t'i-piperazinedione precursor compound was isolated, the following procedure was initiated.

Through a mechanically stirred suspension of 29.95g (0.l7mol) of 2,2,5,5-tetramethyl-3,-piperazinedione in 250ml of water at C, chlorine gas was bubbled therethrough for a period ranging from 1.5 to 2.0 hours. A white solid was isolated by filtration, washed thoroughly with cold water, and dried in vacuo over calcium sulfate to yield 38.20g (0.l6mol) of 1,4- dichloro-2,2,5,5-tetramethyl-3,o-piperazinedione, 94% yield,mp l76.5-l78C; I.R. (KBr)-l680cm '(C=O).

Anal. Calculated for C H, Cl N- O was as follows'.C,40.18;I-l,5.06;N, l I72; and Cl,29.66. Found:C,40,49; H, 5.18; N,ll.64 and Cl,28.7l.

ANTIBACTERIAL ACTIVITY STUDIES Overni ht Organism ATCC Broth ulture Code (Organisms/ml) Staph. epidermidis l2228 X l0 E. coli 10536 l0 X [0' Kleb. pneumoniae l003l 12 X l0'- 3 X l0 Pseud. aeruginosa 9027 I2 X l0- 13 X l0 Staph. aureus 6538 6 X l0 8 X l0 Bord. bronchiseptica 4617 3 X [0 Nutrient Broth B.B.L. 8g/l000 ml. distilled water. The broth contains 5g. gelysate peptone and 3g. beef extract. The solution has a pH of 6.9.

Nutrient Agar 23g/l000 ml. distilled water. The

nutrient contains 5g. gelysate peptone, 3g. beef extract and 15g. agar. Horse Serum T.C. 10% horse serum solution in A distilled water. The serum solution was freshly prepared and adjusted to a pH of 7 using carbon dioxide prior to its use.

METHOD A stock solution of the isolated compound l,4,dichloro-2,2,5,5-tetramethyl-3,-piperazinedione) was prepared using an appropriate buffered solution.

For screening in the absence of a denaturing agent (e.g., horse serum) an equal volume of distilled water and the resulting solution was subjected to the screen.

For screening in the presence of a denaturing agent, a volume of the stock solution was diluted using an equal volume of 10% horse serum. When necessary, the final solution was adjusted to the desired pH using IN HCL and the solution was permitted to stand at room temperature for 30 minutes prior to the screening procedure.

To 5ml. of the stock solution being evaluated, there was added 0.2ml. of an overnight broth culture containing the particular micro-organism being investigated (see above). At time intervals of 0.5, l, 2, 3, 4, 5, minutes, a loop of this suspension was subcultured into 5ml. of'a sterile nutrient broth. All the samples were then incubated at 37C for 7 days with daily observation for evidence of bacterial growth. The time interval reported is for that sample in which no bacterial growth was observed after the incubation period.

Aside from the foregoing, several controls were also employed as described below.

CONTROL 1 This control was designed basically to insure viability of the overnight broth culture.

To 5ml. of a sterile 0.9% sodium chloride solution, there was added 0.2ml. of an overnight broth culture containing the particular micro-organism being investigated. A loop of this suspension was subcultured into 5ml. of a sterile nutrient broth and incubated at 37C for 7 days.

CONTROL 2 This control was designed to insure that the dilution factor of the nutrient broth was beyond any bacteriostatic activity of the l,4-dichloro-2,2,5,S-tetramethyl- 3,6-piperazinedione.

To 5ml of a sterile nutrient broth, there was added a loop of the l,4-dichloro-2,2,5,5-tetramethyl-3,6-piperazinedione solution and the solution was mixed immediately. To this solution, there was then added a loop of an overnight broth culture which was diluted 25x with a 0.9% sodium chloride solution. Incubation was carried out for 7 days at a temperature of 37C.

CONTROL 3 This control was employed to insure the bacterial growth observed was that due to the organism being tested, rather than contamination from a foreign organism.

At the same time intervals used for subculturing the test solution into nutrient broth during the screening procedure, a loop of the test solution was also subcultured onto sterile nutrient agar plates. This technique was useful for observing the characteristic colonial morphology of each organism.

CONTROL 4 TABLE ll-(ontinued This control was used initially to insure that the pH R, R2 R1 I of the solution and the concentration of the buffer species did not inhibit the bacterial growth during the time 5 CH3 (32H:i c n CnH" intervals used in the screening procedure. n lr a u 1 3 1.

2H5 CH3 u n s u The entire screening procedure was conducted for CHH CHn H3 H5 each buffered solution using the buffered solution 2 5 u n u za is rn rather than the 1,4-dichloro-2,2.S,5-tetramethyl-3,6- 5 :1 5:11 5: 1;; 5:53 piperazinedione solution in the procedure. C m. C I-l C,H C,.l-l The results of the antibacterial studies respective of 3 5 n-* 'r" i,4-dichloro-2,2,5,S-tetramethyl-3 6-piperazinedione 3 i i; are set out on the followlng page in Table l. ll fl a n w m I ddf s d d 'b d I B II II N IQ HI RII H na ition, e o owing compoun sas escn e in CNN CmHm Cu",a 5"" "I able ll below, will exhibit substantially, the same antl- 15 u Q Semi i gins!" bacterial activity observed for the compound 1,4- Q Q gfigfi Z Q fi fi dichloro-2,2,5,5-tetramethyl-3,fi-pr erazmedione. NAPHTHYL NAPHTHYL NAPHTHYL NAPHTHYL NAPHTHYL PHENYL NAPHTHYL PHENYL These compounds can be prepared in accordance with PHENYL NAPHTHYL PHENYL NAPHTHYL those procedures previously outlined.

1- A a L s COMPOUND CONDITIONS CONCENTRATION mm mrrucrenm. AC'l'lVl'lY TIME urn u :DILUEN'! cnmounn Positive e1 1222: 1055s 1003;. 9021 053a 4617 Q a I 1 1 l. 1 i -*H 0.1M NaOAc H2O 11.511 x 1e' i4 41-m n 2 1 z s s 1 a c4 13116.6 1.59 m

" 0.1M NOAC Serum 0.59m 10'3M' 1 41 PPM :15 10 10 10 15 10 PH4.6 159m! 2 4 1-1 0.52 x 1o' u 36 m4 2 1 4 s 4 s 0.1m Nat-1 90 se um 0.52 x 34 g 56 PPM 15 15 15 15 15 pH7.0 -1 62Pu. mmu n o, 11 9 'omgxltdy'fu 43 m4 2 4 1 4 4 4 4 pH8.B 4pm:

TABLE 11 STABILITY STUDIES The stability of the compound, l,4-dichloro-2,2,5,S- I R2 R3 R.

tetramethyl-3,6-piperazined1one in aqueous solution cm, (3H,, (3,, qu was determined by (1) following the disappearance of g g" 2 g n" gi the positive chlorine concentration of the N- ih'l. ZLHZ. ci ril, 1 Chloramine solution iodometrically and/or 2 followmfi Crag CHE c n ing the decrease in the absorbance at the maximum z fi h: 45 wavelength spectrophotometrically. cl-i PHENYL CH3 IP'HENYL The data obtained was interpreted in terms of a first g n- 8's order kinetic process and the half-life of said com- Z I; M; 41 pound was reported in units of time at the condition gul s! I" 5.0: I: under which they were determined.

dz The half-life studies between the compound 1.4- C, u C H dichloro-2,2,5,5-tetramethyl-3,6-piperazinedione and 5:13 APHTHYL 8' KJAPHWYL the prior art compound, l,4-dichloro-2,S-piperazined- 4,, CHu tone (per Kokorudz) are found in Table III reproduced gt i v h -L1 below.

H 2 N As is readily apparent, at the same pH (4.6.7.0 and a":- Cu s, 9.3), a compound of this invention. 1,4-dichloro- QQ CHa gfigfi CH 2,2,5,S-tetramethyl-Z. piperazinedione) exhibited far I: C,H,, c,H, greater stability (half-life) than the compound of the g g Elfin prior art (l,4-dichloro,2,5-piperazinedione.) r d; il 0 As a result thereof, the compounds of this invention If gas 1: 811: are far more suitable for commercial application, as 2 3 aqueous solutions of such compounds can remain starJAPHTHYL v CH, vAPH'rI-IYL cu, ble over much longer periods of time, whereas an aqueous solution of the prior art compound would have to c,,|-r,, c rbe employed for antibacterial purpose rapidly, in order g -:4 that antibacterial activity can be exhibited before deg- 3 H 3 radation or decomposition of the compound occurs. m 4| Gu 4. With the compounds of the present invention, stable swam ANALrsIs F QONDI'IIIONS w W TmPERATuRE HAtF-LIFEm I 0' 0.1mm? no 930 g it- L6- k omnanPo io P l 1 1 Er?? .1 10 tmgao aqueous solutions are provided, thus permitting the user to employ such solutions for achieving an antibacterial effect over a much longer period of time than that under which aqueous solutions of the prior art compounds could be employed.

As with the antibacterial activity studies, the compounds set out in Table II, will also exhibit similar stability values as exhibited by l,4-dichloro-2,5-piperazinedione (using the same procedure outlined above) As indicated earlier, the uniqueness of the present invention resides in the fact that the compounds disclosed herein exhibit enhanced stability in aqueous solution over the closest related prior art compounds and in addition, the compounds of this invention exhibit effective antibacterial activity as well. The key to the enhanced stability exhibited by the compounds of the present invention resides in the fact that no alphahydrogen atoms are maintained on the carbon atom adjacent the carbonyl group of the piperazinedione nu- 45 cleus.

As any skilled artisan will understand, the solubility of the compounds of the present invention, in aqueous solution will vary, depending upon the chain length of the alkyl group and further depending upon whether an aryl moiety is introduced. However, any skilled artisan can recognize that for those compounds encompassed within applicants generic formula, which exhibits limited water solubility, such problems can easily be overcome by introducing a suitable conventional solubilizing agent into the aqueous system. As an example, any surfactant, compatible with the N-Chloro functionality, will suffice for applicants purpose.

These compounds are conveniently used in aqueous c1- N-Cl wherein R R,, R, and R, are the same or different, and represent alkyl of from 1 to 20 carbon atoms, phenyl or naphthyl.

2. The compound of claim 1: l,4-diehloro-2,2,5,5-tetramethyl-3,6-piperazinedione. 3. The compound of claim 1: l ,4-dichloro-2,2,5 ,5-tetraethyl-3 ,fi-piperazinedione. 4. The compound of claim 1: l,4-dichloro-2,2,5 ,5-tetrapropyl-3,6-piperazinedione 5. The compound of claim 1: l,4-dichloro-2,2,5,5-tetrabutyl-3,6-piperazinedione 6. The compound of claim 1: l ,4-dichloro,2,2,5 ,5-tetrapentyl-3 ,-piperazinednone.

i l I k 

1. A COMPOUND HAVING THE FOLLOWING FORMULA:
 2. The compound of claim 1: 1,4-dichloro-2,2,5,5-tetramethyl-3,6-piperazinedione.
 3. The compound of claim 1: 1,4-dichloro-2,2,5,5-tetraethyl-3,6-piperazinedione.
 4. The compound of claim 1: 1,4-dichloro-2,2,5,5-tetrapropyl-3,6-piperazinedione
 5. The compound of claim 1: 1,4-dichloro-2,2,5,5-tetrabutyl-3,6-piperazinedione
 6. The compound of claim 1: 1,4-dichloro,2,2,5,5-tetrapentyl-3,6-piperazinedione. 